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(41-14) Flow cytometric analysis of the oxidative stress induced by herbicides as a microalgal cytotoxicity marker. Rioboo, Carmen*,1, O'Connor, Enrique2, Herrera, Guadalupe2, González, Óscar1, Herrero, Concepción1, Cid, Ángeles1, 1 Lab. Microbioloxía,Dpto. Bioloxía Celular e Molecular, Universidade da Coruña, A Coruña, Spain2 Centro de Citiometría, Dpto. Bioquímica y Biología Molecular, Universidad de Valencia, Valencia, Spain ABSTRACT- The herbicide contamination of surface waters has a potential toxicological risk on aquatic environments. Unicellular microalgae can respond inmediatly to environmental changes and, since they are the major primary producers in some aquatic environments could report disturbances at higher levels in the food web, which generally react on longer time scales. Oxidative stress has become one of the main cell issues in cytotoxicity process and its study requires highly sensitive techniques for detection and quantification, since most reactive oxygen species (ROS) have a very short half life but a wide range of biological effects. In this way, we have applied the flow cytometric technique to assess the oxidative activity induced in the freshwater microalga Chlorella vulgaris by the acute exposure at two wide-spread used herbicides, paraquat and terbutryn. Paraquat (a bypyridilic herbicide) is a known donor of O2.- prooxidant; free radical scavengers, such as microalgal carotenoids, are rapidly overwhelmed due to the efficiency of this herbicide in generating free radicals. Terbutryn (a triazine herbicide) is a photosynthetic inhibitor, but no data exist on its potential oxidative toxicity. The microalgal oxidative status was assessed by measuring intracellular levels of superoxide ion with dihydroethidine, a colourless fluorochrome which is oxidized to ethidium by this oxygenic species; then ethidium intercalates within cell DNA, staining its nucleus with a bright fluorescent orange (605 nm). Results obtained showed that both assayed herbicides provoked an increase in the intracellular O2.- generation related with an increased cytometric fluorescence. Moreover, changes in microalgal O2.- levels can provide an early indication of potential problems from contamination with a variety of toxicants and can be a reliable marker of cytotoxicity. PGIDT01MAM10302PR Key words: flow cytometry, herbicides, microalgae, oxidative stress (cytotoxiciy marker) |
